While Na+, K+-ATPase activity has previously been found to be sensitive to ethanol, the role of this integral membrane enzyme as an effector of the neurophysiological actions of ethanol is poorly understood. We have determined that a component of Na+, K+-ATPase-dependent synaptosomal K+ uptake, which is defined as very sensitive (VS; IC50 less than or equal to 10-7M) to inhibition by ouabain, is selectively activated by ethanol (EC50=3.3mM) under apparent resting conditions. Following increased intracellular Na+ (i.e., depolarization), however, basal VS activity increases and the response to ethanol shifts to inhibition. The depolarization-dependent sensitization of Na+, K+-ATPase activity to inhibition (IC25=10 mM) by ethanol is transient and parallels a sensitization to inhibition by very low concentrations (IC25=10-12 M) of ouabain. Importantly, these concentrations of ouabain are far below the concentration (10-8 M) of Na+, K+-ATPase enzymes in the experimental preparation. Recent work suggests that the basis for this apparent sub- stoichiometric inhibition of Na+, K+-ATPase activity[unreadable]by ouabain results fro a protein kinase A-dependent coupling of ouabain binding to the activation of arachidonic acid metabolism and the production of a secondary inhibitor. We are currently elucidating the precise nature of this putative pathway for the amplification of Na+, K+- ATPase inhibition by ouabain and will determine the relevance of this pathway to inhibition by ethanol.